The H-NOX protein structure adapts to different mechanisms in sensors interacting with nitric oxide.

Some classes of bacteria within phyla possess protein sensors identified as homologous to the heme domain of soluble guanylate cyclase, the mammalian NO-receptor. Named H-NOX domain (Heme-Nitric Oxide or OXygen-binding), their heme binds nitric oxide (NO) and O for some of them. The signaling pathways where these proteins act as NO or O sensors appear various and are fully established for only some species.

Ultrafast dynamics of heme distortion in the O-sensor of a thermophilic anaerobe bacterium.

Heme-Nitric oxide and Oxygen binding protein domains (H-NOX) are found in signaling pathways of both prokaryotes and eukaryotes and share sequence homology with soluble guanylate cyclase, the mammalian NO receptor. In bacteria, H-NOX is associated with kinase or methyl accepting chemotaxis domains. In the O-sensor of the strict anaerobe Caldanaerobacter tengcongensis (Ct H-NOX) the heme appears highly distorted after O binding, but the role of heme distortion in allosteric transitions was not yet evidenced.

Soluble Guanylate Cyclase Inhibitors Discovered among Natural Compounds.

Soluble guanylate cyclase (sGC) is the human receptor of nitric oxide (NO) in numerous kinds of cells and produces the second messenger 3',5'-cyclic guanosine monophosphate (cGMP) upon NO binding to its heme. sGC is involved in many cell signaling pathways both under healthy conditions and under pathological conditions, such as angiogenesis associated with tumor growth. Addressing the selective inhibition of the NO/cGMP pathway is a strategy worthwhile to be investigated for slowing down tumoral angiogenesis or for curing vasoplegia.

Multicolor two-photon imaging of endogenous fluorophores in living tissues by wavelength mixing.

Two-photon imaging of endogenous fluorescence can provide physiological and metabolic information from intact tissues. However, simultaneous imaging of multiple intrinsic fluorophores, such as nicotinamide adenine dinucleotide(phosphate) (NAD(P)H), flavin adenine dinucleotide (FAD) and retinoids in living systems is generally hampered by sequential multi-wavelength excitation resulting in motion artifacts. Here, we report on efficient and simultaneous multicolor two-photon excitation of endogenous fluorophores with absorption spectra spanning the 750-1040 nm range, using wavelength mixing.

The Dynamics Behind the Affinity: Controlling Heme-Gas Affinity via Geminate Recombination and Heme Propionate Conformation in the NO Carrier Cytochrome c'.

Nitric oxide (NO) sensors are heme proteins which may also bind CO and O. Control of heme-gas affinity and their discrimination are achieved by the structural properties and reactivity of the heme and its distal and proximal environments, leading to several energy barriers. In the bacterial NO sensor cytochrome c' from Alcaligenes xylosoxidans (AXCP), the single Leu16Ala distal mutation boosts the affinity for gas ligands by a remarkable 10-10-fold, transforming AXCP from one of the lowest affinity gas binding proteins to one of the highest.

Targeting of Helicobacter pylori thymidylate synthase ThyX by non-mitotoxic hydroxy-naphthoquinones.

ThyX is an essential thymidylate synthase that is mechanistically and structurally unrelated to the functionally analogous human enzyme, thus providing means for selective inhibition of bacterial growth. To identify novel compounds with anti-bacterial activity against the human pathogenic bacterium Helicobacter pylori, based on our earlier biochemical and structural analyses, we designed a series of eighteen 2-hydroxy-1,4-naphthoquinones (2-OH-1,4-NQs) that target HpThyX. Our lead-like molecules markedly inhibited the NADPH oxidation and 2'-deoxythymidine-5'-monophosphate-forming activities of HpThyX enzyme in vitro, with inhibitory constants in the low nanomolar range.

Motion of proximal histidine and structural allosteric transition in soluble guanylate cyclase.

We investigated the changes of heme coordination in purified soluble guanylate cyclase (sGC) by time-resolved spectroscopy in a time range encompassing 11 orders of magnitude (from 1 ps to 0.2 s). After dissociation, NO either recombines geminately to the 4-coordinate (4c) heme (τG1 = 7.

Substrate interaction dynamics and oxygen control in the active site of thymidylate synthase ThyX.

Thymidylate synthase ThyX, required for DNA synthesis in many pathogenic bacteria, is considered a promising antimicrobial target. It binds FAD and three substrates, producing dTMP (2'-deoxythymidine-5'-monophosphate) from dUMP (2'-deoxyuridine-5'-monophosphate). However, ThyX proteins also act as NADPH oxidase by reacting directly with O2.

Reactivity and dynamics of H2S, NO, and O2 interacting with hemoglobins from Lucina pectinata.

Hemoglobin HbI from the clam Lucina pectinata is involved in H2S transport, whereas homologous heme protein HbII/III is involved in O2 metabolism. Despite similar tertiary structures, HbI and HbII/III exhibit very different reactivity toward heme ligands H2S, O2, and NO. To investigate this reactivity at the heme level, we measured the dynamics of ligand interaction by time-resolved absorption spectroscopy in the picosecond to nanosecond time range.

Picosecond binding of the His ligand to four-coordinate heme in cytochrome c': a one-way gate for releasing proximal NO.

We provide a direct demonstration of a "kinetic trap" mechanism in the proximal 5-coordinate heme-nitrosyl complex (5c-NO) of cytochrome c' from Alcaligenes xylosoxidans (AXCP) in which picosecond rebinding of the endogenous His ligand following heme-NO dissociation acts as a one-way gate for the release of proximal NO into solution. This demonstration is based upon picosecond transient absorption changes following NO photodissociation of the proximal 5c-NO AXCP complex. We have determined the absolute transient absorption spectrum of 4-coordinate ferrous heme to which NO rebinds with a time constant τ(NO) = 7 ps (k(NO) = 1.

Picosecond to second dynamics reveals a structural transition in Clostridium botulinum NO-sensor triggered by the activator BAY-41-2272.

Soluble guanylate cyclase (sGC) is the mammalian endogenous nitric oxide (NO) receptor that synthesizes cGMP upon NO activation. In synergy with the artificial allosteric effector BAY 41-2272 (a lead compound for drug design in cardiovascular treatment), sGC can also be activated by carbon monoxide (CO), but the structural basis for this synergistic effect are unknown. We recorded in the unusually broad time range from 1 ps to 1 s the dynamics of the interaction of CO binding to full length sGC, to the isolated sGC heme domain β(1)(200) and to the homologous bacterial NO-sensor from Clostridium botulinum.

Absorption band III kinetics probe the picosecond heme iron motion triggered by nitric oxide binding to hemoglobin and myoglobin.

To study the ultrafast movement of the heme iron induced by nitric oxide (NO) binding to hemoglobin (Hb) and myoglobin (Mb), we probed the picosecond spectral evolution of absorption band III (∼760 nm) and vibrational modes (iron-histidine stretching, ν(4) and ν(7) in-plane modes) in time-resolved resonance Raman spectra. The time constants of band III intensity kinetics induced by NO rebinding (25 ps for hemoglobin and 40 ps for myoglobin) are larger than in Soret bands and Q-bands. Band III intensity kinetics is retarded with respect to NO rebinding to Hb and to Mb.

In vivo structural imaging of the cornea by polarization-resolved second harmonic microscopy.

The transparency and mechanical strength of the cornea are related to the highly organized three-dimensional distribution of collagen fibrils. It is of great interest to develop specific and contrasted in vivo imaging tools to probe these collagenous structures, which is not available yet. Second Harmonic Generation (SHG) microscopy is a unique tool to reveal fibrillar collagen within unstained tissues, but backward SHG images of cornea fail to reveal any spatial features due to the nanometric diameter of stromal collagen fibrils.

Quaternary structure controls ligand dynamics in soluble guanylate cyclase.

Soluble guanylate cyclase (sGC) is the mammalian endogenous nitric oxide (NO) receptor. The mechanisms of activation and deactivation of this heterodimeric enzyme are unknown. For deciphering them, functional domains can be overexpressed.

The anti-Müllerian hormone type II receptor: insights into the binding domains recognized by a monoclonal antibody and the natural ligand.

Anti-Müllerian hormone (AMH) [also called Müllerian inhibiting substance (MIS)] is a member of the transforming growth factor-beta family. AMH and its type II receptor (AMHR-II) are involved in the regression of the Müllerian ducts in the male embryo, and in gonadal functions in the adult. AMH is also known to be a marker of granulosa and Sertoli cell tumours.

Engagement of bone morphogenetic protein type IB receptor and Smad1 signaling by anti-Müllerian hormone and its type II receptor.

Anti-Müllerian hormone induces the regression of fetal Müllerian ducts and inhibits the transcription of gonadal steroidogenic enzymes. It belongs to the transforming growth factor-beta family whose members signal through a pair of serine/threonine kinase receptors and Smad effectors. Only the anti-Müllerian hormone type II receptor has been identified.

Reduction of insulin and triglycerides delays glomerulosclerosis in obese Zucker rats.

To evaluate the effect of insulin and/or triglycerides on the pathogenesis of glomerulosclerosis, acarbose (BAYg5421), an inhibitor of intestinal alpha-glucosidases, was administered as a dietary admix (40 mg/100 g chow) to Zucker obese rats (ZOA), from 1.5 months until sacrifice at 1.5, 5, 8, 10 and 15 months.

Insensitivity to anti-müllerian hormone due to a mutation in the human anti-müllerian hormone receptor.

Anti-Müllerian hormone (AMH) and its receptor are involved in the regression of Müllerian ducts in male fetuses. We have now cloned and mapped the human AMH receptor gene and provide genetic proof that it is required for AMH signalling, by identifying a mutation in the AMH receptor in a patient with persistent Müllerian duct syndrome. The mutation destroys the invariant dinucleotide at the 5' end of the second intron, generating two abnormal mRNAs, one missing the second exon, required for ligand binding, and the other incorporating the first 12 bases of the second intron.

Anti-müllerian hormone in early human development.

Anti-müllerian hormone (AMH) is a glycoprotein produced by immature Sertoli cells and responsible for the regression of müllerian ducts in male fetuses. The ontogeny of the hormone in early human development was investigated. While no detectable AMH could be found in female fetal serum, in males, the mean +/- S.